Glial cells in the gut represent the morphological and functional equivalent of astrocytes and microglia in the central nervous system (CNS). In recent years, the role of enteric glial cells (EGCs) has extended from that of simple nutritive support for enteric neurons to that of being pivotal participants in the regulation of inflammatory events in the gut. Similar to the CNS astrocytes, the EGCs physiologically express the S100B protein that exerts either trophic or toxic effects depending on its concentration in the extracellular milieu. In the CNS, S100B overexpression is responsible for the initiation of a gliotic reaction by the release of pro-inflammatory mediators, which may have a deleterious effect on neighboring cells. S100B-mediated pro-inflammatory effects are not limited to the brain: S100B overexpression is associated with the onset and maintenance of inflammation in the human gut too. In this review we describe the major features of EGCs and S100B protein occurring in intestinal inflammation deriving from such.

Intestinal tissues are innervated by a complex and extensive component known as the enteric nervous system (ENS)[1]. The ENS is characterized by the presence of neurons and enteric glial cells (EGCs) which are arranged into interconnected ganglia distributed between 2 major plexuses, and they control several gut functions[2,3]. During the course of time, the traditional view of EGCs has changed from being a mere mechanical support for surrounding neurons to that of a more articulate and complex nature, since they are actively involved in the regulation of homeostasis, motility and inflammatory processes within the gut[4,5].

Similar to the astrocytes in the central nervous system (CNS), the EGCs release several signaling molecules[4,5]. Among these, great importance has been given to the better comprehension of the specific glial-derived S100B protein[6-8]. This protein is a small, diffusible neurotrophin that is situated in the cytoplasm and/or the nucleus of both nervous and non-nervous tissues[9,10]. In the brain, S100B has been considered a “Janus face” neurotrophin[11,12] because it exerts opposite actions depending on its concentration in the extracellular milieu: it has a pro-proliferative and neurogenic effect on astroglia and on serotonergic neurons at nanomolar concentrations, as well as a neurodegenerative function[12] at micromolar concentrations, determining mysregulated glial cell proliferation and amplifying neuroinflammation. Similar to the brain, recent studies have suggested the involvement of S100B in inflammatory processes occurring in the gut, highlighting the importance of EGCs as key regulators of gut homeostasis[13-15].

In this review, we will focus on the role of EGCs and the S100B protein and we will take them into consideration by looking at both experimental animal models and some human diseases for which evidence exists, and in particular their involvement in inflammatory conditions of the human gut where the role of S100B appears to be prominent.

The gut is characterized by a sequence, starting from the serosa, as follows: subserosa, longitudinal muscle, myenteric plexus, circular muscle, submucosal plexus, muscularis mucosae and mucosa[2]. The myenteric and submucosal plexuses are characterized by the presence of ganglia which, in turn, contain enteric neurons and EGCs in a ratio of 1:7[16]. In the ENS, the enteric ganglia are involved in basic gut functions, such as the regulation of peristalsis, secretion and blood flow and the modulation of the immune/inflammatory processes[17-19]. Several neurotransmitters are involved in the control of all these intestinal functions, such as vasoactive intestinal peptide and nitric oxide (NO)[20,21]. In particular, NO is produced by the biosynthetic enzyme neuronal NO synthase (NOS), which is expressed in myenteric neurons, and by the inducible form of NOS (iNOS), which is expressed in EGCs[15].

In humans, recent and increasing data has demonstrated that EGCs and S100B protein are directly involved in gut inflammatory diseases[13,14,33]. Previous investigations described abnormalities of the enteroglial network in patients with Crohn’s disease and UC[33]. More recently, glial abnormalities have been confirmed by 2 separate studies carried out by our group[13,14]. In particular, we demonstrated that, in patients with celiac disease (CD)[13] and UC[14], EGCs participate in the modulation of mucosal NO production via S100B overexpression and release. Indeed, in patients with CD, we demonstrated that S100B plays an active role in NO-dependent inflammation[13]. In particular, increased S100B protein expression and release were observed in the duodenal mucosa of patients with untreated CD, compared to healthy controls (Figure 1A)[13]. Very interestingly, S100B upregulation was accompanied by enhanced iNOS protein expression and consequent NO release, both representing crucial features in CD[62].

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Figure 1 Changes in S100B protein expression during intestinal inflammation. A: Celiac disease[13]. Immunohistochemistry shows stronger S100B immunopositivity in the duodenal mucosa of patients affected by celiac disease, compared with healthy controls (original magnification, × 100). The graphs represent S100B protein expression (left) and release (right) in healthy controls and patients with celiac disease (^bP < 0.01); B: Ulcerative colitis[14]. Immunohistochemistry shows stronger S100B immunopositivity in the rectal submucosa of patients with ulcerative colitis, compared with healthy controls (original magnification, × 100). The graphs represent S100B protein expression (left) and release (right) in healthy controls and patients with ulcerative colitis (^dP < 0.01).

The relationship between S100B and NO production was confirmed by the demonstration that the administration of exogenous S100B protein to non-inflamed duodenal biopsy specimens from healthy controls, resulted in both iNOS protein expression and NO release, indicating that micromolar concentrations of this protein are able to participate in the inflammatory response of even “healthy duodenum”[13]. Besides NO production, exogenous S100B mediates a significant increase in lipid peroxidation associated with a marked increase in phosphorylated-p38 MAPK protein expression and with the activation of NF-κB, in accordance with the previously mentioned studies[13].

Taken together, these observations represented the first data in humans suggesting that via S100B upregulation, EGCs directly participate in NO-dependent inflammation occurring in CD, and they paved the way by supposing that EGCs are part of complex immuno-regulatory effectors since they establish a strategic first defense line against foreign antigens. By means of EGC proliferation, changes in enteroglial architecture have been reported also in patients with IBD[33,63]. Several studies have shown that EGC markers are differentially altered in Crohn’s disease and UC with a decrease in EGC density in Crohn’s disease and a gliosis-like phenomenon in UC[33,63].

In support of these observations, it has recently been confirmed that EGCs directly participate in the chronic mucosal inflammation of patients with UC[14]. In fact, S100B immunoreactivity significantly increased in the rectal mucosa of these patients when compared to the mucosal S100B expression in healthy controls (Figure 1B)[14]. This upregulation was associated with the specific stimulation of iNOS and consequent abnormal mucosal NO production, both representing characteristic features of UC[64,65].

In addition, via iNOS expression, exogenous S100B induces a significant and concentration-dependent increase in NO production in the human rectal mucosa of healthy controls via RAGE interaction[14], confirming the ability of EGCs to modulate NO production and the specificity of S100B protein-mediated responses in the human gut.

Further confirming that EGCs are part of the complex system of immunoregulatory effectors in the gut, it has been shown that the addition of pro-inflammatory stimuli to rectal mucosal tissue led to EGC activation, again via RAGE involvement[14] as demonstrated by both S100B upregulation and enhanced NO production. These findings indicate that EGCs are able to recognize inflammatory stimuli and that once activated, they produce and release S100B up to micromolar concentrations, thereby contributing to NO production in the human gut.